It is widely accepted that intrathecal injection of opioids and alpha2 adrenergic drugs produce antinociception in animal models of pain and in man. The sites and mechanism of action of these drugs at the level of the spinal cord, however remains unknown. one possible mechanism to explain this spinal action of opioids and alpha2-agonists is by inhibition of neurotransmitter release from nociceptive sensory neurons. The purpose of this work is to determine whether tolerance and cross-tolerance develop to the inhibitory effects of opioids and alpha2-agonists on transmitter release from sensory neurons. Two techniques will be used to study tolerance and cross-tolerance, release evoked from spinal cord slices and release from rat sensory neurons. Regulation of release of substance P and calcitonin gene-related peptide are the endpoints of the studies. The use of spinal cord slices will allow correlation between tolerance to antinociception and tolerance to inhibition of transmitter release. Use of sensory neurons in culture will provide a model for studying tolerance mechanisms. Regulation of potassium-stimulated release will be measured in spinal cords from non-tolerant rats and rats tolerant to the antinociceptive effects of opioids or alpha2 agonists. Dose-response curves for inhibition of release will be generated for both groups and will be compared to determine if tolerance and cross-tolerance develop. Similar experiments will be performed in rat sensory grown in cells in grown culture. Regulation of release of peptides in cells chronically exposed to morphine will be compared with naive cells. In addition, selective studies will be performed on cells in culture to determine if G-proteins and/or cyclic AMP are involved in acute regulation of transmitter release and in the development of tolerance. From these studies we will gain knowledge as to the sites and mechanisms by which opioids and alpha2-agonists produce antinociception and on the mechanisms of tolerance development.